Research in our group explores the mechanisms of genome stability and the consequences for cancer, including the biochemical mechanism of repair of cross-links between DNA strands and the DNA polymerases that help cells tolerate DNA damage.
This area of our research examines the biochemical mechanism of repair of cross-links between DNA strands. Chemicals that produce cross-links are particularly destructive to genome function and thus, are widely used in cancer chemotherapy.
Exogenous sources of intra-stand crosslinking chemicals include mitomycin C, nitrogen mustard, cisplatin, and psoralen. Endogenous sources creating crosslinks include metabolites created by lipid peroxidation such as acrolein, crotonaldehyde, malondialdehyde and nitric oxide. These agents are widely used for the treatment of leukemia and solid tumors. However, the mechanism of repair in mammalian cells is not well understood and the nucleotide excision repair (NER) pathway may be highly involved in this process. We study how NER proteins help repair DNA crosslinks.
1P01 CA193124 NIH/NCI “Processing and Repair of DNA Crosslinks”
(Wood, PI) 02/01/2017 – 01/31/2022
Manandhar M, Lowery MG, Boulware KS, Lin KH, Lu Y, Wood RD. (2017) Transcriptional consequences of XPA disruption in human cell lines. DNA Repair. 57:76-90.
DAPI stained micronuclei
DNA Polymerase theta (POLQ)
We have also identified and are studying the function of other DNA processing enzymes. We have initially cloned the mammalian gene and identified some of the functions of POLQ. This specialized DNA polymerase is important for the repair of DNA double strand-breaks (DSBs) from many sources including x-rays and cancer chemotherapy drugs.
The image to the left shows micronuclei, products from unrepaired DSBs. The frequency of cells displaying micronuclei is higher in POLQ deficient cells than in normal cells.
RP150102 CPRIT “GenomeStability and Immune Diversity Controlled by the POLQ Pathway” (McBride, PI; Wood Co-PI) 03/01/2015-2/28/2018
Yousefzadeh MJ, Wyatt DW, Takata K, Mu Y, Hensley SC, Tomida J, Bylund GO, Doublié S, Johansson E, Ramsden DA, McBride KM, Wood RD. (2014) Mechanism of suppression of chromosomal instability by DNA polymerase POLQ. PLoS Genetics, 10:10 e1004654 doi: 10.1371/journal.pgen.1004654
REV3L knockout mouse skin epithelium
DNA polymerase zeta (pol ζ) is exceptionally important for controlling mutagenesis and genetic instability in cells. REV3L is the catalytic subunit of pol ζ, while REV7 (MAD2L2) is considered an accessory subunit.
We have generated mouse models with specific deficiencies in DNA polymerase zeta. These models reveal that the enzyme is critical for maintaining chromosome stability and limiting tumorigenesis. Deletion of Rev3L in p53 mutant mice leads to increased cancer incidence and a shortened lifespan. Further, two distinct REV7-binding sites in REV3L are needed to prevent chromosome breaks and confer resistance to DNA damage.
W81XWH-17-1-0239 Department of Defense (CDMRP) DoD Prostate Cancer Idea Development Award, “DNA polymerase zeta inactivationin prostate cancer” (Wood, PI) 06/15/2017 – 06/14/2019
1R03 CA212556-01 NIH/NCI “Loss of novel end joining factor with dysfunction of BRCA1 contributes to treatment resistance” (Tomida, PI) 08/08/2017 – 07/31/2019
MD Anderson Cancer Center Prostate Cancer SPORE Developmental Research Program “Identifying DNA polymerase zeta inactivationin prostate cancer to improve therapy” 03/01/2017 – 02/28/2018
Lange SS, Tomida J, Boulware KS, Bhetawal S, Wood RD. (2016) The Polymerase Activity of Mammalian DNA Pol ζ Is Specifically Required for Cell and Embryonic Viability. PLoS Genet.12(1):e1005759
Section of testis from POLN deficient mouse
We have discovered POLN as a homologue of POLQ are working to identify its biochemical activities. The POLN gene is predominantly expressed in
testis and encodes protein with strand displacement and damage bypass activity.
The image to the right shows gamma-H2AX foci (green dots, places of DNA damage) induced by x-ray irradiation in spermatid cells lacking POLN.
Takata KI, Reh S, Yousefzadeh MJ, Zelazowski MJ, Chetawal S, Trono D, Lowery MG, Sandoval M, Takata Y, Lu Y, Lin K, Shen J, Kusewitt DF, McBride KM, Cole F Wood RD. (2017) Analysis of DNA polymerase ν function in meiotic recombination, immunoglobulin class-switching, and DNA damage tolerance. PLoS Genetics 13(6):e1006818
DNA Helicase HELQ
Agents that cause interstrand DNA crosslinks (ICL) are widely used to treat cancer. Our work found that the DNA helicase HELQ helps defend human cells against ICL- inducing agents, associating with ATR-ATRIP DNA damage signaling and homologous recombination components such as BCDX2 that are sometimes altered in ovarian cancer
In the image, the red jagged line outlined in gray represents an interstrand crosslink. HELQ along with ATR and RAD51 paralogs help cells tolerate such interstrand crosslinks.
Takata K, Reh S, Tomida J, Person MD, Wood RD. (2013). Human DNA helicase HELQ participates in DNA interstrand crosslink tolerance with ATR and RAD51 paralogs. Nature Communications 4:2338.